Abstract
Ykt6 is a soluble N-ethylmaleimide sensitive factor activating protein receptor (SNARE) critically involved in diverse vesicular fusion pathways. While most SNAREs rely on transmembrane domains for their activity, Ykt6 dynamically cycles between the cytosol and membrane-bound compartments where it is active. The mechanism that regulates these transitions and allows Ykt6 to achieve specificity toward vesicular pathways is unknown. Using a Parkinson's disease (PD) model, we found that Ykt6 is phosphorylated at an evolutionarily conserved site which is regulated by Ca2+ signaling. Through a multidisciplinary approach, we show that phosphorylation triggers a conformational change that allows Ykt6 to switch from a closed cytosolic to an open membrane-bound form. In the phosphorylated open form, the spectrum of protein interactions changes, leading to defects in both the secretory and autophagy pathways, enhancing toxicity in PD models. Our studies reveal a mechanism by which Ykt6 conformation and activity are regulated with potential implications for PD.
Highlights
Ykt6 is a soluble N-ethylmaleimide sensitive factor activating protein receptor (SNARE) critically involved in diverse vesicular fusion pathways
Ykt6 plays a key role in numerous vesicular transport pathways in yeast and in mammalian cells: 1) the secretory pathways, which are endoplasmic reticulum (ER) to the Golgi apparatus (Golgi), intra-Golgi, Golgi–ER retrieval pathways, and in the constitutive transport from the Golgi to the plasma membrane [3, 5, 6]; 2) the endocytic pathways, which are between the Golgi and the vacuole/ lysosome [7, 8] and endosomes to exosomes [9]; and 3) the macroautophagy pathway [10,11,12]
Through a multidisciplinary approach encompassing both in vitro and in vivo analyses in yeast, worms, and human cells, we reveal a regulatory mechanism by which Ykt6 operates
Summary
Ykt is a soluble N-ethylmaleimide sensitive factor activating protein receptor (SNARE) critically involved in diverse vesicular fusion pathways. The mechanism that regulates these transitions and allows Ykt to achieve specificity toward vesicular pathways is unknown. We show that phosphorylation triggers a conformational change that allows Ykt to switch from a closed cytosolic to an open membrane-bound form. The reversible nature of palmitoylation and/or geranylgeranylation has been proposed as a mechanism to regulate Ykt membrane association, allowing it to cycle between the cytosol and membrane-bound compartments [14, 17]. We report that phosphorylation regulated by Ca2+ signaling drives a conformational change that allows Ykt to switch from a closed cytosolic to an open membrane-bound form. Phosphorylation is a critical determinant for Ykt protein interactions with functional consequences in the secretory and autophagy pathways under normal and α-synuclein conditions. Whether Ykt regulation by phosphorylation contributes to Ykt deficits under α-syn toxicity remains to be determined
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